体外共培养研究乳腺癌细胞株MCF-7对正常内皮细胞的作用

Influence of breast carcinoma cells on normal endothelial cells: experimental study with a co-culture system

目的在体外共培养体系中研究乳腺癌细胞对正常内皮细胞的作用。方法建立乳腺癌细胞株MCF7与正常内皮细胞的体外共培养体系条件培养液ZMCF7EC;以同期单独培养的正常内皮细胞为对照,对与乳腺癌细胞株MCF7在ZMCF7EC条件下共培养后的内皮细胞进行透射电镜细胞超微结构、光镜细胞形态学观察,并以看家基因β2m为内参照进行ESM、IGFBP3、αvβ3、VEC、及Tie22基因半定量RTPCR表达分析,以缩时录像技术进行血管新生观察。结果与同期单独培养的正常内皮细胞对照相比,经共培养后的内皮细胞形态不规则、呈游走状;超微结构出现胞核增大、核仁增大、核浆比例增大,内质网疏松、变形,细胞表面窗孔加深,细胞间隙加大和细胞表面纤毛减少;半定量RTPCR值均较对照上调,其半定量值与对照组比较:ESM/β2m(0.60±0.28,P<0.01)、IGFBP3/β2m(0.54±0.30,P<0.05)、αvβ3/β2m(0.58±0.23,P<0.01)、VEC/β2m(0.51±0.17,P<0.01)及Tie22/β2m(0.55±0.21,P<0.05)。结论共培养后的内皮细胞有明显的趋瘤特征和血管新生能力,推测活体内乳腺癌生长中通过对相邻内皮细胞的作用促进血管新生且新生血管的内皮细胞性质与行为不同于正常内皮细胞。

Objective To study the influence of breast carcinoma cells on normal endothelial cells. Methods Human endothelial cells were isolated from umbilical cord blood. Medium Z-MCF-7-EC was established and was used to co-culture the normal endothelium cells (ECs) and human breast cancer cells of the line MCF-7. Normal endothelial cells cultured by itself were used as control. Light microscopy and transmission electron microscopy were used to observe the morphology of the 2 kinds of endothelial cells. Expression of the genes ESM, IGFBP-3, αvβ3, VE-C, and Tie-2-2 was analyzed by semi-quantitative RT-PCR using a house-keeping gene β2m as inner reference gene. Results The ECs co-cultured with MCF-7breast cancer cells were abnormal in shape with increased size of nucleus and nucleolus, increased size ratio of nuclear to nucleoplasm, increased depth of surface fenestration, loosed and distorted endoplasm, increased size of cell-cell junctions, decreased number of surface microvilli, and tubules formed by ECs. The expression values of the genes ESM, IGFBP-3, αvβ3, VE-C, and Tie-2-2 in the ECs co-cultured with MCF-7 breast cancer cells were all up-regulated in comparison with those in the controls (P<0.01, P<0.05,P<0.01,P<0.01, and P<0.05 respectively). Conclusion Breast cancer cells promote formation of new vessels with endothelial cells different from the normal ECs in character and behavior.